scholarly journals Functions of human replication protein A (RPA): From DNA replication to DNA damage and stress responses

2006 ◽  
Vol 208 (2) ◽  
pp. 267-273 ◽  
Author(s):  
Yue Zou ◽  
Yiyong Liu ◽  
Xiaoming Wu ◽  
Steven M. Shell
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Stress Responses ◽  
Protein A ◽  
Replication Protein A ◽  
Replication Protein

scholarly journals DNA Replication but Not Nucleotide Excision Repair Is Required for UVC-Induced Replication Protein A Phosphorylation in Mammalian Cells

2000 ◽  
Vol 20 (8) ◽  
pp. 2696-2705 ◽  
Author(s):  
Gregory Rodrigo ◽  
Sophie Roumagnac ◽  
Marc S. Wold ◽  
Bernard Salles ◽  
Patrick Calsou
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Protein A ◽  
Replication Protein A ◽  
Replication Protein ◽  
Dependent Manner ◽  
Nucleotide Excision

ABSTRACT Exposure of mammalian cells to short-wavelength light (UVC) triggers a global response which can either counteract the deleterious effect of DNA damage by enabling DNA repair or lead to apoptosis. Several stress-activated protein kinases participate in this response, making phosphorylation a strong candidate for being involved in regulating the cellular damage response. One factor that is phosphorylated in a UVC-dependent manner is the 32-kDa subunit of the single-stranded DNA-binding replication protein A (RPA32). RPA is required for major cellular processes like DNA replication, and removal of DNA damage by nucleotide excision repair (NER). In this study we examined the signal which triggers RPA32 hyperphosphorylation following UVC irradiation in human cells. Hyperphosphorylation of RPA was observed in cells from patients with either NER or transcription-coupled repair (TCR) deficiency (A, C, and G complementation groups of xeroderma pigmentosum and A and B groups of Cockayne syndrome, respectively). This exclude both NER intermediates and TCR as essential signals for RPA hyperphosphorylation. However, we have observed that UV-sensitive cells deficient in NER and TCR require lower doses of UV irradiation to induce RPA32 hyperphosphorylation than normal cells, indicating that persistent unrepaired lesions contribute to RPA phosphorylation. Finally, the results of UVC irradiation experiments on nonreplicating cells and S-phase-synchronized cells emphasize a major role for DNA replication arrest in the presence of UVC lesions in RPA UVC-induced hyperphosphorylation in mammalian cells.

scholarly journals Adozelesin Triggers DNA Damage Response Pathways and Arrests SV40 DNA Replication through Replication Protein A Inactivation

2000 ◽  
Vol 275 (2) ◽  
pp. 1391-1397 ◽  
Author(s):  
Jen-Sing Liu ◽  
Shu-Ru Kuo ◽  
Mary M. McHugh ◽  
Terry A. Beerman ◽  
Thomas Melendy
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Dna Damage Response ◽  
Protein A ◽  
Replication Protein A ◽  
Replication Protein ◽  
Damage Response ◽  
Sv40 Dna

scholarly journals Focus-formation of replication protein A, activation of checkpoint system and DNA repair synthesis induced by DNA double-strand breaks in Xenopus egg extract

2002 ◽  
Vol 115 (15) ◽  
pp. 3159-3169 ◽  
Author(s):  
Takayuki Kobayashi ◽  
Shusuke Tada ◽  
Takashi Tsuyama ◽  
Hiromu Murofushi ◽  
Masayuki Seki ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Replication ◽  
Protein A ◽  
Replication Protein A ◽  
Replication Protein ◽  
Focus Formation ◽  
Xenopus Egg Extract ◽  
Egg Extract ◽  
Xenopus Egg

The response to DNA damage was analyzed using a cell-free system consisting of Xenopus egg extract and demembranated sperm nuclei. In the absence of DNA-damaging agents, detergent-resistant accumulation of replication protein A appeared in nuclei after a 30 minute incubation, and a considerable portion of the replication protein A signals disappeared during a further 30 minute incubation. Similar replication protein A accumulation was observed in the nuclei after a 30 minute incubation in the extract containing camptothecin, whereas a further 30 minute incubation generated discrete replication protein A foci. The addition of camptothecin also induced formation of γ-H2AX foci, which have been previously shown to localize at sites of DSBs. Analysis of the time course of DNA replication and results obtained using geminin, an inhibitor of licensing for DNA replication, suggest that the discrete replication protein A foci formed in response to camptothecin-induced DNA damage occur in a DNA-replication-dependent manner. When the nuclei were incubated in the extract containing EcoRI,discrete replication protein A foci were observed at 30 minutes as well as at 60 and 90 minutes after incubation, and the focus-formation of replication protein A was not sensitive to geminin. DNA replication was almost completely inhibited in the presence of EcoRI and the inhibition was sensitive to caffeine, an inhibitor of ataxia telangiectasia mutated protein (ATM) and ATM- and Rad3-related protein (ATR). However, the focus-formation of replication protein A in the presence of EcoRI was not influenced by caffeine treatment. EcoRI-induced incorporation of biotin-dUTP into chromatin was observed following geminin-mediated inhibition of DNA replication, suggesting that the incorporation was the result of DNA repair. The biotin-dUTP signal co-localized with replication protein A foci and was not significantly suppressed or stimulated by the addition of caffeine.

scholarly journals Replication Protein A (RPA) Phosphorylation Prevents RPA Association with Replication Centers

2004 ◽  
Vol 24 (5) ◽  
pp. 1930-1943 ◽  
Author(s):  
Vitaly M. Vassin ◽  
Marc S. Wold ◽  
James A. Borowiec
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Protein A ◽  
Simian Virus 40 ◽  
Replication Protein A ◽  
Replication Protein ◽  
Wild Type ◽  
Chromosomal Dna

ABSTRACT Mammalian replication protein A (RPA) undergoes DNA damage-dependent phosphorylation at numerous sites on the N terminus of the RPA2 subunit. To understand the functional significance of RPA phosphorylation, we expressed RPA2 variants in which the phosphorylation sites were converted to aspartate (RPA2D) or alanine (RPA2A). Although RPA2D was incorporated into RPA heterotrimers and supported simian virus 40 DNA replication in vitro, the RPA2D mutant was selectively unable to associate with replication centers in vivo. In cells containing greatly reduced levels of endogenous RPA2, RPA2D again did not localize to replication sites, indicating that the defect in supporting chromosomal DNA replication is not due to competition with the wild-type protein. Use of phosphospecific antibodies demonstrated that endogenous hyperphosphorylated RPA behaves similarly to RPA2D. In contrast, under DNA damage or replication stress conditions, RPA2D, like RPA2A and wild-type RPA2, was competent to associate with DNA damage foci as determined by colocalization with γ-H2AX. We conclude that RPA2 phosphorylation prevents RPA association with replication centers in vivo and potentially serves as a marker for sites of DNA damage.

Replication Protein A phosphorylation and the cellular response to DNA damage

DNA Repair ◽  
2004 ◽  
Vol 3 (8-9) ◽  
pp. 1015-1024 ◽  
Author(s):  
Sara K. Binz ◽  
Anne M. Sheehan ◽  
Marc S. Wold
Keyword(s):  
Dna Damage ◽  
Cellular Response ◽  
Protein A ◽  
Replication Protein A ◽  
Replication Protein
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